Structural Biochemistry/Proteins/Analysis of Purification Result
After each purification steps, the types of protein that exist in the in the solution is expected to decrease while its specific activity is expected to increase. These two qualities are desirable because experiment done using a pure protein sample gives a more quantifiable result. One method used to check the purity of the sample is using a form of Gel Electrophoresis, such as SDS PAGE or native PAGE.
Purification can also be quantitatively evaluated by measuring total protein, total activity, specific activity, yield and purification level. Total protein is the quantity of protein present in a fraction and can be determined by measuring the protein concentration of a part of each fraction and multiplying by the fraction's total volume. Total activity is measured by the enzymatic activity in the volume of fraction used in the assay multiplied by the fraction's total volume. Specific activity is the total activity divided by total protein. The yield is the amount of activity retained after each purification step. The purification level is the increase of purity which can be measured after each purification step by dividing its specific activity by the specific activity of the initial extract.
a good purification takes into account both purification levels of yield. A high amount of purification and a poor yield give little protein to work with. on the other hands, a low purification and a high yield give contaminated protein in the experiment.